We have profiled gene expression in mouse skin to identify those genes that are regulated in a stage specific manner during the hair cycle. About third of skin-expressed genes show changes in expression in relation to the hair growth cycle. These genes can be grouped into distinct clusters based on the expression pattern. To begin understanding how hair follicle cells are specified, we propose to focus on delineating the mechanisms underlying expression of markers of hair follicle keratinocyte terminal differentiation, which are predominantly hair keratin and keratin associated proteins. Based on the time-course expression data, we have identified several clusters containing the terminal differentiation genes and a set of transcription factors, which we propose are critical for hair follicle specification. Our hypothesis is that these transcription factors associate with the regulatory regions encoding the terminal differentiation markers, as well as affecting the expression of each other. We now wish to use new genome-based approaches to understand in greater detail the transcriptional networks underlying hair follicle specification. To establish the feasibility of our approach, we initially plan to focus on a set of transcription factors for which there is significant experimental data in support of their importance in hair follicle specification. The specific aims are to: 1. Create a promoter/tiling microarray for selective hair follicle differentiation genes, which can be used to map transcription factor binding sites. 2. Use the ChlP-chip method to define binding sites of key transcription factors to regulatory regions of genes encoding hair differentiation markers and transcription factors. 3. Improve computational methods to determine binding sites for key transcription factors in hair follicle differentiation genes. Interpreted in combination with gene expression data and analyses of mice with mutations in these factors, we expect to build a model to describe the series of gene expression events involved in construction of the hair follicle. [unreadable] [unreadable]